The present invention relates to the use of leptin antagonists for treating insulin resistance in Type II diabetes and to a pharmaceutical for treating such resistance.
Diabetes is one of the most frequently occurring metabolic diseases in industrialized countries. There are some 110 million diabetics world-wide; while approx. 10 million of these are Type I diabetics, the overwhelming majority (approx. 100 million) are Type II diabetics. The disease is caused by faulty regulation of glucose metabolism. In Type I diabetes, failure of the xcex2 cells in the pancreas results in insulin no longer being formed. This lack of insulin leads to an increase in blood glucose and, if not treated by supplying insulin, to ketoacidosis, diabetic coma and death of the patient. In Type II diabetics, the causal relationships are different and are characterized by the initial development of insulin resistance, i.e. diminution in the ability of the cells to respond adequately to insulin. Excessive weight and lack of physical activity, in particular, are regarded as being responsible for inducing insulin resistance. The latter condition is not noticed initially since it is offset by an increased secretion of insulin. However, the continuing insulin resistance leads, in a process extending over many years, to failure of the endogenous compensation mechanism and consequent development of Type II diabetes. While diet and physical activity can delay this sequence of events, they are frequently unable to prevent manifestation of the disease. Medicinal intervention is then required in order to control the blood glucose adequately.
It is of crucial importance for the long-term success of the therapy that the blood glucose be maintained as narrowly as possible within the physiological range. It is the current view that glucose levels which have been elevated for decades, as are found in poorly controlled diabetics (both Type I and Type II diabetics), make an important contribution to late complications in diabetes. These late complications constitute, in particular, blood vessel damage which leads to kidney diseases, loss of sight and cardiovascular diseases. This so-called late damage is an important factor contributing to mortality in diabetics.
In 1994, a new hormone, leptin, was described which is formed in fat cells and which is lacking in genetically overweight mice (ob/ob mice) (Zhang, Y., Proenca, R., Maffei, M., Barone, M., Leopold, L., and Friedman, J. M. (1994). Positional cloning of the mouse obese gene and its human homologue. Nature 372, 425-432.). Human leptin and murine leptin are to a large extent identical. Injecting ob/ob mice with recombinantly prepared leptin leads to a reduction in nutrient intake and to a decrease in weight (Pelleymounter, M. A., Cullen, M. J., Baker, M. B., Hecht, R., Winters, D., Boone, T., and Collins, F. (1995). Effects of the obese gene product on body weight regulation in ob/ob mice. Science 269, 540-543.). There has so far been no indication that mutations in the ob gene might be responsible for the frequent occurrence of obesity in humans (approx. 30% of the population is markedly overweight in the USA). Systematic investigations have demonstrated that serum levels of leptin are increased in obese humans as they are in various animal models of obesity (Dagogo-Jack, S., Fanelli, C., Paramore,: D., Brothers, J., and Landt, M. (1996). Plasma leptin and insulin relationships in obese and nonobese humans. Diabetes 45, 695-698; Considine, R. V., Sinha, M. K., Heiman, M. L., Kriauciunas, A., Stephens, T. W., Nyce, M. R., Ohannesian, J. P., Marco, C. C., McKee, L. J., Bauer, T. L., and Caro, J. F. (1996). Serum immunoreactive leptin concentrations in normal-weight and obese humans. N. Engl. J. Med. 334, 292-295.). For this reason, it is assumed that leptin is a feedback signal which informs the brain of the quantity of energy which is stored in the fat tissue. According to this assumption, it is then the function of the brain to decrease feed intake by inhibiting appetite, on the one hand, and to stimulate basal metabolism on the other. In human obesity, this regulatory circuit appears to be interrupted.
In addition to this, it is assumed that leptin also acts directly on tissues outside the brain.
Three studies relating to the direct effect of leptin on isolated cells have so far been published:
Kroder et al. (Kroder, G., Kellerer, M., and Hxc3xa4ring, H. (1996) Exp. Crin. Endocrin. Diabetes 104 Suppl. 2, 66 (Abstract)) made the assumption that leptin establishes a connection between insulin resistance and obesity and report that leptin decreases insulin-induced phosphorylation of the insulin receptor and of insulin receptor substrate 1 (IRS-1) in rat 1 fibroblasts which are overexpressing the human insulin receptor. The extent to which leptin also exerts an influence on the end points of the insulin effect, for example stimulation of glucose transport or glycogen synthase, was not investigated or discussed.
It has been demonstrated that sensitivity to lipogenic hormones (dexamethasone and insulin) is decreased in transformed 30A5 preadipocytes which are overexpressing leptin (Bai, Y. L., Zhang, S. Y., Kim, K. S., Lee, J. K., and Kim, K. H. (1996) J. Biol. Chem. 271, 13939-13942). Fatty acid synthesis and the synthesis of neutral lipids were decreased by leptin overexpression even in the non-stimulated state. While control cells exhibited a marked increase in the rate of lipid synthesis after the cells had been treated With dexamethasone or insulin, or a combination of the two hormones, cells which were overexpressing leptin were hardly stimulated at all under these circumstances. In addition to this, an investigation was also undertaken of the inhibition of glycerophosphate dehydrogenase activity and of acetyl CoA carboxylase expression which occurs after treating the cells with a combination of dexamethasone and insulin. It was found that it was not possible to stimulate leptin-expressing cells. The effects which were observed indicate that leptin suppresses lipid metabolism in a general manner. There is no mention of any possible connection between obesity and insulin resistance.
In another model system, i.e. C2C12 mouse myotubes, it was found that leptin exhibits insulin-like effects (Berti, L., Kellerer, M., and Hxc3xa4ring, H. (1996) Diabetologia 39 Suppl. 1, A59 (Abstract)). This study reports that both glucose transport and glycogen synthesis are stimulated by leptin. These findings conflict with those reported by the other authors and the results which are presented here. They possibly involve an effect which is specific for this cell type.
In our investigations into the effect of leptin on isolated rat adipocytes, a model system for fat tissue, it has now been found, surprisingly, that the insulin sensitivity of important metabolic pathways of the fat cell such as stimulation of lipogenesis, of glucose transport and of glycogenesis, is drastically reduced (Example 4) whereas the basal values remain unaffected. The same applies to the inhibition of isoproterenol-stimulated lipolysis. Glucose transport in isolated rat adipocytes is stimulated approximately 14-fold by adding insulin (10 nM). This capacity to be stimulated is reduced in a dose-dependent manner by preincubating with leptin at different concentrations for 15 hours. Leptin desensitizes the cells, i.e. an insulin resistance is produced. Dose/effect curves for insulin at different leptin concentrations (Example 5) demonstrate that the concentrations at which effects can already be detected in vitro, both as regards insulin (0.1-0.2 nM) and as regards leptin (0.5-1 nM), are within the physiological range (Dagogo-Jack et al., 1996; Considine et al., 1996). Higher leptin levels (2-4 nM) (Dagogo-Jack et al., 1996; Considine et al., 1996), are found in obese humans, ,so that it is possible that the insulin effect is more strongly impaired in these subjects. The conclusion therefore suggests itself that chronically elevated leptin, as can be seen in obese subjects, leads to insulin resistance. As already explained above, insulin resistance is an important factor in the pathogenesis of Type II diabetes.
It is therefore an object of the invention to provide novel leptin antagonists that may be formulated in pharmaceutical compositions. It is another object of the invention to provide methods of treating Type II diabetes and other insulin-related disorders.
The invention consequently relates to the use of leptin antagonists, in particular those which are derived from leptin itself, for preparing a pharmaceutical for use in Type II diabetes. The leptin antagonists for this use are described in more detail below.
According to a first object of the invention, pharmaceutical compositions are provided which comprise an antagonist of leptin. According to this same object, pharmaceutical compositions are disclosed which comprise leptin antagonsists which are derived from leptin. Further according to this object, pharmaceutical compositions are revealed which comprise a leptin antagonist that is a soluble leptin receptor, or a derivative thereof.
According to a second object of the invention, methods are provided which utilize the inventive pharmaceutical compositions in the treatment of Type II diabetes. Also according to this object, methods are provided for restoring or amplifying the physiological effect of insulin.